The proposed study will investigate regulation of the insulin receptor (IR) by insulin and glucocorticoids. When in excess both hormones can cause insulin resistance and diabetes. The first series of studies will focus on the in vitro regulation of the IR by glucocorticoids. We have recently reported that, in IM-9 cultured lymphocytes, glucocorticoids increase IR mRNA levels and IR gene transcription. In addition, we have sequenced the promoter region of the IR gene, and have identified a potential glucocorticoid response element (GRE) consensus sequence in the first intron. DNA binding, DNAase I sensitivity and transient (and/or stable) transfection studies will be carried out to determine whether this sequence functions as a GRE to convey glucocorticoid regulation upon the IR. Further genomic sequences will be obtained to identify additional regulatory elements. The second series of studies will focus on the in vitro regulation of the IR by insulin. In several cell types we have recently observed that insulin decreases IR biosynthesis and mRNA levels. We plan to determine whether this effect occurs either at the level of RNA transcription or at the level of mRNA stability. If the effect occurs at the level of transcription, then the regulatory elements in the IR gene will be investigated. Parallel to these two in vitro studies, in vivo studies will be carried out in adrenalectomized, sham operated and corticosterone-treated normal and diabetic rats. Since in vivo studies indicate that glucocorticoids down-regulate the IR whereas most in vitro studies indicate that glucocorticoids up-regulate the IR, we will attempt to explain the variables occurring in vivo. One hypothesis is that the hyperinsulinemia induced by glucocorticoids in vivo down- regulates the IR and overrides the up-regulation induced by glucocorticoids. These in vitro and in vivo studies should provide important information, therefore, concerning the mechanisms through which two major hormones regulate the IR.